Kevin Michael Dalton

All Publications

• Scaling and merging time-resolved pink-beam diffraction with variational inference STRUCTURAL DYNAMICS-US Zielinski, K. A., Dolamore, C., Wang, H. K., Henning, R. W., Wilson, M. A., Pollack, L., Srajer, V., Hekstra, D. R., Dalton, K. M. 2024; 11 (6)

  View details for DOI 10.1063/4.0000269

  View details for Web of Science ID 001351080800002

• Laue-DIALS: Open-source software for polychromatic x-ray diffraction data STRUCTURAL DYNAMICS-US Hewitt, R. A., Dalton, K. M., Mendez, D. A., Wang, H. K., Klureza, M. A., Brookner, D. E., Greisman, J. B., McDonagh, D., Srajer, V., Sauter, N. K., Brewster, A. S., Hekstra, D. R. 2024; 11 (5): 054701

  Abstract

  Most x-ray sources are inherently polychromatic. Polychromatic ("pink") x-rays provide an efficient way to conduct diffraction experiments as many more photons can be used and large regions of reciprocal space can be probed without sample rotation during exposure-ideal conditions for time-resolved applications. Analysis of such data is complicated, however, causing most x-ray facilities to discard >99% of x-ray photons to obtain monochromatic data. Key challenges in analyzing polychromatic diffraction data include lattice searching, indexing and wavelength assignment, correction of measured intensities for wavelength-dependent effects, and deconvolution of harmonics. We recently described an algorithm, Careless, that can perform harmonic deconvolution and correct measured intensities for variation in wavelength when presented with integrated diffraction intensities and assigned wavelengths. Here, we present Laue-DIALS, an open-source software pipeline that indexes and integrates polychromatic diffraction data. Laue-DIALS is based on the dxtbx toolbox, which supports the DIALS software commonly used to process monochromatic data. As such, Laue-DIALS provides many of the same advantages: an open-source, modular, and extensible architecture, providing a robust basis for future development. We present benchmark results showing that Laue-DIALS, together with Careless, provides a suitable approach to the analysis of polychromatic diffraction data, including for time-resolved applications.

  View details for DOI 10.1063/4.0000265

  View details for Web of Science ID 001326831800001

  View details for PubMedID 39386198

  View details for PubMedCentralID PMC11462730

• Scaling and Merging Time-Resolved Laue Data with Variational Inference. bioRxiv : the preprint server for biology Zielinski, K. A., Dolamore, C., Wang, H. K., Henning, R. W., Wilson, M. A., Pollack, L., Srajer, V., Hekstra, D. R., Dalton, K. M. 2024

  Abstract

  Time-resolved X-ray crystallography (TR-X) at synchrotrons and free electron lasers is a promising technique for recording dynamics of molecules at atomic resolution. While experimental methods for TR-X have proliferated and matured, data analysis is often difficult. Extracting small, time-dependent changes in signal is frequently a bottleneck for practitioners. Recent work demonstrated this challenge can be addressed when merging redundant observations by a statistical technique known as variational inference (VI). However, the variational approach to time-resolved data analysis requires identification of successful hyperparameters in order to optimally extract signal. In this case study, we present a successful application of VI to time-resolved changes in an enzyme, DJ-1, upon mixing with a substrate molecule, methylglyoxal. We present a strategy to extract high signal-to-noise changes in electron density from these data. Furthermore, we conduct an ablation study, in which we systematically remove one hyperparameter at a time to demonstrate the impact of each hyperparameter choice on the success of our model. We expect this case study will serve as a practical example for how others may deploy VI in order to analyze their time-resolved diffraction data.

  View details for DOI 10.1101/2024.07.30.605871

  View details for PubMedID 39131362

• Sensitive Detection of Structural Differences using a Statistical Framework for Comparative Crystallography. bioRxiv : the preprint server for biology Hekstra, D. R., Wang, H. K., Klureza, M. A., Greisman, J. B., Dalton, K. M. 2024

  Abstract

  Chemical and conformational changes underlie the functional cycles of proteins. Comparative crystallography can reveal these changes over time, over ligands, and over chemical and physical perturbations in atomic detail. A key difficulty, however, is that the resulting observations must be placed on the same scale by correcting for experimental factors. We recently introduced a Bayesian framework for correcting (scaling) X-ray diffraction data by combining deep learning with statistical priors informed by crystallographic theory. To scale comparative crystallography data, we here combine this framework with a multivariate statistical theory of comparative crystallography. By doing so, we find strong improvements in the detection of protein dynamics, element-specific anomalous signal, and the binding of drug fragments.

  View details for DOI 10.1101/2024.07.22.604476

  View details for PubMedID 39091831

• Resolving DJ-1 Glyoxalase Catalysis Using Mix-and-Inject Serial Crystallography at a Synchrotron. bioRxiv : the preprint server for biology Zielinski, K. A., Dolamore, C., Dalton, K. M., Smith, N., Termini, J., Henning, R., Srajer, V., Hekstra, D. R., Pollack, L., Wilson, M. A. 2024

  Abstract

  DJ-1 (PARK7) is an intensively studied protein whose cytoprotective activities are dysregulated in multiple diseases. DJ-1 has been reported as having two distinct enzymatic activities in defense against reactive carbonyl species that are difficult to distinguish in conventional biochemical experiments. Here, we establish the mechanism of DJ-1 using a synchrotron-compatible version of mix-and-inject-serial crystallography (MISC), which was previously performed only at XFELs, to directly observe DJ-1 catalysis. We designed and used new diffusive mixers to collect time-resolved Laue diffraction data of DJ-1 catalysis at a pink beam synchrotron beamline. Analysis of structurally similar methylglyoxal-derived intermediates formed through the DJ-1 catalytic cycle shows that the enzyme catalyzes nearly two turnovers in the crystal and defines key aspects of its glyoxalase mechanism. In addition, DJ-1 shows allosteric communication between a distal site at the dimer interface and the active site that changes during catalysis. Our results rule out the widely cited deglycase mechanism for DJ-1 action and provide an explanation for how DJ-1 produces L-lactate with high chiral purity.

  View details for DOI 10.1101/2024.07.19.604369

  View details for PubMedID 39071394

• Resolving conformational changes that mediate a two-step catalytic mechanism in a model enzyme. bioRxiv : the preprint server for biology Greisman, J. B., Dalton, K. M., Brookner, D. E., Klureza, M. A., Sheehan, C. J., Kim, I., Henning, R. W., Russi, S., Hekstra, D. R. 2023

  Abstract

  Enzymes catalyze biochemical reactions through precise positioning of substrates, cofactors, and amino acids to modulate the transition-state free energy. However, the role of conformational dynamics remains poorly understood due to lack of experimental access. This shortcoming is evident with E. coli dihydrofolate reductase (DHFR), a model system for the role of protein dynamics in catalysis, for which it is unknown how the enzyme regulates the different active site environments required to facilitate proton and hydride transfer. Here, we present ligand-, temperature-, and electric-field-based perturbations during X-ray diffraction experiments that enable identification of coupled conformational changes in DHFR. We identify a global hinge motion and local networks of structural rearrangements that are engaged by substrate protonation to regulate solvent access and promote efficient catalysis. The resulting mechanism shows that DHFR's two-step catalytic mechanism is guided by a dynamic free energy landscape responsive to the state of the substrate.

  View details for DOI 10.1101/2023.06.02.543507

  View details for PubMedID 37398233

• An information theoretic framework reveals a tunable allosteric network in group II chaperonins. Nature structural & molecular biology Lopez, T., Dalton, K., Tomlinson, A., Pande, V., Frydman, J. 2017; 24 (9): 726-733

  Abstract

  ATP-dependent allosteric regulation of the ring-shaped group II chaperonins remains ill defined, in part because their complex oligomeric topology has limited the success of structural techniques in suggesting allosteric determinants. Further, their high sequence conservation has hindered the prediction of allosteric networks using mathematical covariation approaches. Here, we develop an information theoretic strategy that is robust to residue conservation and apply it to group II chaperonins. We identify a contiguous network of covarying residues that connects all nucleotide-binding pockets within each chaperonin ring. An interfacial residue between the networks of neighboring subunits controls positive cooperativity by communicating nucleotide occupancy within each ring. Strikingly, chaperonin allostery is tunable through single mutations at this position. Naturally occurring variants at this position that double the extent of positive cooperativity are less prevalent in nature. We propose that being less cooperative than attainable allows chaperonins to support robust folding over a wider range of metabolic conditions.

  View details for DOI 10.1038/nsmb.3440

  View details for PubMedID 28741612

  View details for PubMedCentralID PMC5986071

• The Mechanism and Function of Group II Chaperonins. Journal of molecular biology Lopez, T., Dalton, K., Frydman, J. 2015; 427 (18): 2919-2930

  Abstract

  Protein folding in the cell requires the assistance of enzymes collectively called chaperones. Among these, the chaperonins are 1-MDa ring-shaped oligomeric complexes that bind unfolded polypeptides and promote their folding within an isolated chamber in an ATP-dependent manner. Group II chaperonins, found in archaea and eukaryotes, contain a built-in lid that opens and closes over the central chamber. In eukaryotes, the chaperonin TRiC/CCT is hetero-oligomeric, consisting of two stacked rings of eight paralogous subunits each. TRiC facilitates folding of approximately 10% of the eukaryotic proteome, including many cytoskeletal components and cell cycle regulators. Folding of many cellular substrates of TRiC cannot be assisted by any other chaperone. A complete structural and mechanistic understanding of this highly conserved and essential chaperonin remains elusive. However, recent work is beginning to shed light on key aspects of chaperonin function and how their unique properties underlie their contribution to maintaining cellular proteostasis.

  View details for DOI 10.1016/j.jmb.2015.04.013

  View details for PubMedID 25936650

• The Dynamic Conformational Cycle of the Group I Chaperonin C-Termini Revealed via Molecular Dynamics Simulation PLOS ONE Dalton, K. M., Frydman, J., Pande, V. S. 2015; 10 (3)

  Abstract

  Chaperonins are large ring shaped oligomers that facilitate protein folding by encapsulation within a central cavity. All chaperonins possess flexible C-termini which protrude from the equatorial domain of each subunit into the central cavity. Biochemical evidence suggests that the termini play an important role in the allosteric regulation of the ATPase cycle, in substrate folding and in complex assembly and stability. Despite the tremendous wealth of structural data available for numerous orthologous chaperonins, little structural information is available regarding the residues within the C-terminus. Herein, molecular dynamics simulations are presented which localize the termini throughout the nucleotide cycle of the group I chaperonin, GroE, from Escherichia coli. The simulation results predict that the termini undergo a heretofore unappreciated conformational cycle which is coupled to the nucleotide state of the enzyme. As such, these results have profound implications for the mechanism by which GroE utilizes nucleotide and folds client proteins.

  View details for DOI 10.1371/journal.pone.0117724

  View details for Web of Science ID 000352134700014

  View details for PubMedID 25822285

  View details for PubMedCentralID PMC4379175

• The dynamic conformational cycle of the group I chaperonin C-termini revealed via molecular dynamics simulation. PloS one Dalton, K. M., Frydman, J., Pande, V. S. 2015; 10 (3)

  Abstract

  Chaperonins are large ring shaped oligomers that facilitate protein folding by encapsulation within a central cavity. All chaperonins possess flexible C-termini which protrude from the equatorial domain of each subunit into the central cavity. Biochemical evidence suggests that the termini play an important role in the allosteric regulation of the ATPase cycle, in substrate folding and in complex assembly and stability. Despite the tremendous wealth of structural data available for numerous orthologous chaperonins, little structural information is available regarding the residues within the C-terminus. Herein, molecular dynamics simulations are presented which localize the termini throughout the nucleotide cycle of the group I chaperonin, GroE, from Escherichia coli. The simulation results predict that the termini undergo a heretofore unappreciated conformational cycle which is coupled to the nucleotide state of the enzyme. As such, these results have profound implications for the mechanism by which GroE utilizes nucleotide and folds client proteins.

  View details for DOI 10.1371/journal.pone.0117724

  View details for PubMedID 25822285

  View details for PubMedCentralID PMC4379175